E1A基因通过抑制NF-κB信号通路增加鼻咽癌细胞放射敏感性实验研究  

Effect of EIA gene on radiosensitivity of human nasopharyngeal carcinoma cells and its possible mechanism

作　　者：肖华平[1] 李庆[1] 谢辉[1] 罗春阳 方玉江[2] 

机构地区：[1]湘南学院附属医院肿瘤科,郴州423000 [2]美国密苏里大学医学院Ellis Fischel肿瘤中心,哥伦比亚65212

出　　处：《中华放射肿瘤学杂志》2017年第11期1327-1331,共5页Chinese Journal of Radiation Oncology

基　　金：湖南省自然科学基金资助项目（14JJ3136）;郴州市科技局资助项目（2012CJ113）

摘　　要：目的 探讨E1A基因对人鼻咽癌细胞放射敏感性的影响及其可能机制。方法 通过腺病毒载体介导,将E1A基因转染至鼻咽癌CNE-2R细胞,采用RT-PCR鉴定E1A基因的表达;分别给予未转染组（PBS组）、转染空载体Ad-β-gal组（Ad-β-gal组）和转染E1A组（Ad-E1A组）的CNE-2R细胞0、2、4、6、8 Gy照射,应用克隆形成实验检测各组CNE-2R细胞放射敏感性的变化;流式细胞术检测各组细胞的凋亡;蛋白印迹法检测各组细胞NF-κB、CK2α、Bcl-2及Cleaved caspase-3蛋白的表达。结果 RT-PCR确认E1A基因已整合到CNE-2R细胞中且稳定表达;Ad-E1A组CNE-2R细胞克隆形成率明显少于PBS和Ad-β-gal组的克隆形成率,Ad-E1A组CNE-2R细胞SF2为0.217小于 PBS组和Ad-β-gal组的0.602和0.585（P〈0.05）,Ad-E1A组α/β值为24.68大于PBS组和Ad-β-gal组的5.268和5.132（P〈0.05）;流式细胞术显示单独放射可促进CNE-2R细胞的凋亡,当与E1A基因联合使用时,细胞凋亡率明显增加（P〈0.05）;蛋白印迹法显示E1A基因可下调NF-κB/P65、CK2α、Bcl-2和上调Cleaved caspase-3的表达。结论 E1A基因可以通过抑制CK2的表达阻断NF-κB信号通路以及促进细胞凋亡,来提高鼻咽癌细胞对放射的敏感性。Objective To investigate the effect of E1A gene on the radiosensitivity of human nasopharyngeal carcinoma cells and its possible mechanism. Methods The E1A gene was transfected into nasopharyngeal carcinoma CNE-2R cells by adenovirus vector. The expression of E1A gene was detected by RT-PCR. Untransfected CNE-2R cells （PBS group） and CNE-2R cells transfected with empty vector Ad-β-gal （Ad-β-gal group） and E1A （Ad-E1A group） were given 0 Gy, 2 Gy, 4 Gy, 6 Gy, 8 Gy 6 MV X-ray irradiation. The changes in radiosensitivity of CNE-2R cells were determined by colony-forming assay. Flow cytometry was used to analyze cell apoptosis in each group. The expression of NF-κB, CK2α, Bcl-2, and cleaved caspase-3 was measured by Western blot. Resutls RT-PCR confirmed that the E1A gene was transfected into CNE-2R cells and stably expressed. The Ad-E1A group had a significantly lower plating efficiency than the PBS group and the Ad-β-gal group （P〈0.05）. The Ad-E1A group had significantly lower cell survival rate at 2 Gy irradiation than the PBS group and the Ad-β-gal group （0.217 vs. 0.602, P〈0.05;0.217 vs. 0.585, P〈0.05）. The Ad-E1A group had a significantly higher α/β value than the PBS group and the Ad-β-gal group （24.680 vs. 5.268, P〈0.05;24.680 vs. 5.132, P〈0.05）. Flow cytometry Resutls showed that irradiation alone could promote the apoptosis of CNE-2R cells, when combined with E1A gene, the apoptosis rate was significantly increased （P〈0.05）. Western blot Resutls showed that E1A gene down-regulated the expression of NF-κB/p65, CK2α, and Bcl-2 and up-regulated the expression of cleaved caspase-3. Conclutions E1A gene can enhance the radiosensitivity of nasopharyngeal carcinoma cells by inhibiting the expression of CK2 to block the NF-κB signaling pathway and promoting cell apoptosis.

关 键 词：E1A基因 核转录因子kappa B 酪蛋白激酶2 鼻咽癌细胞 放射敏感性凋亡 

分 类 号：R739.63[医药卫生—肿瘤]